The objective of this research is to understand the role of the cell surface receptor in generation of the intracellular activator of DNA replication in epidermal growth factor (EGF) induced stimulation of cellular growth. I have developed a photoaffinity labeling technique for specific covalent radiolabeling of the cell surface receptor for EGF. This procedure has been used for studying the dynamic and migratory behavior of the receptor after binding to EGF. This technique will be further exploited to monitor purification of the receptor. Specific antibodies will be generated against the receptor using the hybridoma technique. The anti-receptor antibody will be used as a tool to test the physiological significance of receptor internalization and processing in the biological action of EGF. A cytobiochemical assay will be developed for the EGF-induced intracellular activator of DNA replication. Cytoplasmic extracts from EGF-stimulated cells will be introduced into unstimulated cells by microinjection or encapsulation, and the stimulation of DNA synthesis will be measured. Anti-receptor antibody will be used to deplete the assay system of receptors/processed receptor, and the effects of addition of isolated receptor fragments to these depleted systems will be examined. Properties of the EGF-induced intracellular messenger will be compared with those of the cytoplasmic activator of DNA synthesis in tumor or cancer cells. The cryptic or masked mitogen receptor/processed receptors in transformed cells will be characterized biochemically using anti-receptor antibody. These detailed cytobiochemical studies will aid greatly in achieving my long-term goals: complete definition of the molecular events leading to DNA replication in mitogenic hormone action, and assessment of the roles of mitogenic hormones and their receptors in normal growth and development.